Expression of Notch family proteins in CTCL. (A) Western blot analysis of whole-cell lysates from CTCL cell lines: MyLa (lane 1), SeAx (lane 2), and Hut78 (lane 3). The blots were probed with the antibodies against Notch1 to Notch4; the intracellular domains are visualized as bands at approximately 110 kDa (Notch1 and Notch2), approximately 90 kDa (Notch3), and approximately 52 kDa (Notch4), Notch ligand, Delta, and Hes, the transcriptional factor induced by Notch signaling. (B) Western blot analysis of normal proliferating T cells (P1051) and nonproliferating leukocytes from 2 healthy donors probed for Notch1 to Notch4 and Hes1. (C) Immunostaining for Notch1 in skin biopsies from a patient with tumor-stage MF (top images, UplanSApo 40×/0.90; bottom images, UplanSApo 60×/0.90 using Olympus BX51 microscope with camera Olympus DP70) showing high Notch1 expression in tumor cells. The staining was mainly cytoplasmic and membranous. Arrowhead (top right image) indicates epidermiotropic Notch1-positive tumor cells as displayed in bottom right image at higher magnification. The arrow indicates Notch1-negative small reactive T lymphocytes. (D) Quantification of Notch1 expression in biopsies from lesional skin from patients with MF in stage I (patch and plaque), stage II (tumors), transformed MF, and SS. The histograms represent the percentage of patients classified into one of the 3 categories (negative, moderately positive, and positive), as described in “Immunohistochemistry.” Comparison of the expression profiles shows statistical significant increase in Notch expression with stage (P = .017).